Abstract 18410: Disruption of Beta-Catenin/VEGF/FLK-1/NADPH Oxidase Signaling Leads to Myocardial Ischemia Reperfusion Injury in Ischemic Preconditioned Myocardium: Study With GP91 Knockout Mice
Ischemic preconditioning (IP) mediated activation of VEGF-FLK1-MKK2/NFkappaB pathway identifies these components as a critical regulator of myocardial angiogenesis in an infarcted myocardium. It is well known that reactive oxygen species (ROS) plays an important role in angiogenesis. Out of the several potential sources of ROS in cells, NADPH oxidase is especially important for O2- production. GP91phox is an important regulatory subunit of NADPH oxidase. Hence in the present study we examined the role of ROS and NADPH oxidase in IP mediated myocardial angiogenesis by using gp91Phox−/− knockout (KO) mice. Both wild type (WT) and gp91Phox−/− (GP) mice subjected to either 30 min of I followed by 2 h of reperfusion (R) or IP prior to 30 min I and 2h of R. Reduction in LVDP (62.4 vs. 65.9 mmHg), dp/dtmax (744 vs. 1113 mmHg/sec) and aortic flow (0.9 vs 2.1 ml/min) was observed in GPIPIR compared to WTIPIR along with increased infarct size (31 vs. 21 %) and apoptosis (2.9fold). Illumina gene chip analysis demonstrated 66 (WTIPIR vs. GPIPIR) differentially regulated genes. We further validated selected down regulated genes by real-time RT-PCR such as beta-catenin, SRPK3, ERDR1, ACIN1, Syntaxin-8 and STC1. Among those genes, b-catenin downregulation was thought to be more significant since it was shown to play a role in IP mediated VEGF signaling. To further explore the role of b-catenin in vivo, WT and gp91phox−/− mice were subjected to permanent LAD ligation (MI) with or without IP (4min I and 4min R repeated 3 times). We found increased infarct size in GPIPMI (49%) as compared to WTIPMI (37%) along with decreased capillary and arteriolar density. Echocardiography demonstrated decreased ejection fraction (40 vs. 54 %), fractional shortening (19 vs. 27 %) following 4 weeks of MI in GPIPMI group as compared to WTIPMI. Immunohistochemical analysis showed decrease in nuclear translocation of b-catenin in GPIPMI as compared to WTIPMI. Gel-shift analysis documented reduced NFkappaB DNA binding activity in GPIPMI compared to WTIPMI. Taken together, this study demonstrated for the first time that ROS derived from gp91 containing NADPH oxidase play a critical role in IP mediated myocardial angiogenesis via b-catenin translocation and NFkappaB activation in the infarcted myocardium.
- © 2010 by American Heart Association, Inc.